New improved antibiotics are continually in demand, for the treatment of diseases in man. Antibiotic resistant organisms are continually a problem, with Vancomycin the last defense, particularly in hospitals, vancomycin resistant strains are increasing among the hospital isolated pathogens. A recent survey found 7.9% of Enterococci in the United States hospitals are now vancomycin resistant. (“Nosocomial Enterococci Resistant to Vancomycin” Morbidity and Mortality Weekly Report 42(30):597-598(1993)). Further resistance of Vancomycin and other antibiotics to Enterococcus faecium is reported, (Handwergers. et al., Clin. Infect. Dis. 1993(16),750-755). Resistance organisms are also a problem for other important antibiotics which includes piperacillin. In an effort to overcome resistant organisms and to enhance the effectiveness of antibiotics by inhibiting the activity of a specific enzyme, beta-lactamase, which is produced by certain drug-resistant strains of bacteria, beta-lactamase inhibitors are combined with antibiotics or concurrently administered.
Clearly, antibiotic resistance is a growing public health problem and having new antibiotics available could provide additional options for physicians in treatment regimens. Further, the medical community recognizes that there is an ongoing need for additional antibiotics and concurrently that beta-lactamase inhibitors are useful for enhancing presently available antibiotics to which resistance has developed. The preparation of new beta-lactamase inhibitors requires processes that can efficiently produce intermediates necessary for the preparation of the final beta-lactamase products. As in the case of many synthetic procedures, mixtures of compounds are produced which require chromatographic separations for purification.
The present invention overcomes the problems of purification and separation of mixtures by presenting an alternate method for preparing a single carboxylic acid intermediate through a selective enzymatic hydrolysis of one ester isomer in a mixture of positional ester isomers. Use of hydrolases such as lipases, esterases or proteases for separation of chiral molecules by kinetic resolution is very well established in the literature, (Zaks, A,. Curr. Opinion in Chem Biol. 2001, 5, 130-136.; Wang, C -H and Whitesides G. M. In Enzymes in synthetic organic chemistry: Tetrahedron organic chemistry series vol. 12: pp. 41-130, Pergamon press; Berglund P, and Hult, K. Biocatalytic synthesis of enantiopure compounds using lipases, pp 633-657, In “Stereoselective biocatalysis”, ed. Patel, R. N. Marcel Dekker Inc., 2000). Preferential cleavage by enzymes of one of the ester groups present on the same molecule (also described as regiospecificity) is also known, (Keller, J. W, Hamilton, B. J., Tetrahedron Letts.1986, 27, 1249.; Pirrung M. C., and Krishnamurthy N., J. Org. Chem. 1993, 58, 954).
However, selective hydrolysis of positional ester isomers by enzymatic hydrolysis and separation of the desired resulting carboxylic acid intermediate compounds described in the present invention is not known.